Chemotaxis, directed migration of cells in a gradient of chemoattractant, is an important biological phemonmena that plays vital roles in many physiological processes such as cancer metastasis and wound healing. It is still not clear how the migrating cells inside the tissue find the correct destination while encountering multiple chemoattractant signals in complex spatial and temporal patterns. Despite the importance of chemotaxis in biology, conventional devices and assays for chemotaxis are limited in their ability to generate and maintain complex, stable gradients because they involve point or line sources that produce fluctuating bell-shaped gradients. Efforts to understand the mechanism behind chemotaxis would benefit from a technique that could generate gradients of biologically active chemicals and allow observation of the cellular behavior. The purpose of this proposal is to develop and implement novel microfluidic chemotaxis devices for investigation of neutrophil chemotaxis. We plan to investigate the relative strengths and roles of a range of chemoattractants by exposing the cells to microfluidic concentration gradients of IL-8, MCP-1, and C5a. The microfluidic device will be used to study how the cells migrate to correct targets when presented with two or more competing gradients. The microfluidic chemotaxis chamber will generate complex and overlapping gradients of different soluble factors. Individual cells as well as a population of cells will be followed throughout the experiment while they migrate across a well-defined stable gradient. The use of microfluidic device will help provide clearer understanding of how cells navigate in complex environments in tissues and will find applications in development and testing of new drugs for cancer and wound healing.